Figure 5: Common inversion involving intron 22 of factor 8 gene. A. The F8 gene inversion arises from homologous recombination between int22h-1, a region within intron 22 of the F8 gene, and one of two additional copies of the int22h-1 (a/b) region located approximately 500kb, 5’ and telomeric to the F8 gene. The int22h regions are approximately 9.5kb in length and have 99% homology with one another. Due to intra-chromosomal crossing over between the homologous sequences, an inversion of exons 1-22 away from exonic region 23-26 occurs and disrupts the factor VIII protein. This leads to severe hemophilia A in ~35% to 45% of all cases B. The intron 22 inversion is commonly detected by Long PCR, using two sets of primers specific for the F8 intragenic and extragenic copies and by the differential migration of amplicons during agarose gel electrophoresis. Thus a normal male (-) will exhibit a 10 and 12 Kb amplicon while inversion positive patient will exhibit 10 and 11 kb amplicons. A carrier female (+/-) can be identified by the presence of all the three PCR fragments.